Polymerase chain reaction (“PCR”) is a powerful molecular biology technique used to copy, i.e., amplify, specific nucleic acid sequences within template deoxyribonucleic acid (“DNA”), whether derived from native DNA or cDNA reverse transcribed from ribonucleic acid (“RNA”). An entire PCR assay is carried out in a single tube containing a mixture of enzyme, template, primers, and substrates. Each amplification cycle starts with denaturation (which includes at least heating) that is followed by annealing and then elongation (also known as synthesis and polymerization) reactions.
The PCR assay, in which the cycle of denaturing, annealing and synthesizing reactions is typically repeated 20 or more times, can be divided into three phases: exponential when the PCR reaction product doubles during every cycle (assuming 100% reaction efficiency); linear when the reaction components are consumed, and the reaction product growth slows and degrades; and plateau when the reaction product growth stops and degrades. In traditional (also known as End-Point, or classic) PCR, detection and quantitation of the amplified sequence, i.e., the reaction product, are performed in the plateau phase. In real-time PCR, the amount of PCR reaction product is detected and measured at each cycle during the exponential growth phase, which enables determination of the initial amount of DNA template with great precision.
Enzyme-linked immunosorbent assay (“ELISA”) is a commonly used diagnostic technique, in which an enzyme is coupled directly to an antibody. The antibody bound to an unknown amount of antigen can be quantitated indirectly by measuring the conversion by the enzyme of an ELISA substrate to a product. A detectable signal can permit real-time determination of the ELISA reaction rate, e.g., a color change can be used to quantify enzyme reaction through photometric adsorbency measurement.
Lateral flow technology (“LFT”) assay (also known simply as strip-test) has been a popular platform for diagnostic tests since its introduction in the late 1980s, e.g., the human early pregnancy test. LFT assay has been used for qualitative or semi-quantitative detection of specific analytes including antigens and antibodies, hence other common names for LFT assay: lateral flow immunoassay and immunochromatographic strip test. Biologic sample for LFT assay may be derived from whole blood, serum, plasma, saliva, urine, feces, wound exudate, soil, dust, vegetation, food, or other suitable source. Even products of nucleic acid amplification systems, such as PCR reaction products, can be studied by LFT assay. LFT assay can be run in a single step using the biologic sample in a variety of test locations, such as laboratory, agricultural field, crime scene, etc. Several analytes can be tested simultaneously on the same strip.
The prior art requires a burdensome and time consuming step of eluting (also known as extracting) an analyte from an LFT device before performing the PCR or ELISA technique on the analyte collected by LFT assay. There exists a need in the art for an LFT device configured for collecting (also known as sampling, fixing, or binding) the analyte that may be placed directly in an analysis system for analysis, such as by the PCR or ELISA technique, without the eluting step intervening. Eliminating the eluting step would speed and simplify the collection of specimens for molecular-clinical diagnostics. The apparatus and method embodied by the claims finds application when sampling specimens for molecular-clinical diagnostics in human health, veterinary health, plant health, biosecurity (e.g., surveillance and response), defense, forensics, microbial forensics, and food quality and biosecurity, among other fields of study.